The disc diffusion test is a commonly used, and important, method for determining antibiotic sensitivity of bacteria in the clinical and research laboratory. In this test, paper discs impregnated with different antibiotics are placed onto an agar plate in which bacteria have been spread all over the surface, and the culture is then incubated. If an antibiotic is effective against a particular antibiotic, it will prevent the bacteria from growing, or kill them, so that there will be an area around the disc where the bacteria have not grown. This is called a zone of inhibition, and its appearance indicates sensitivity of the bacterium to the antibiotic. When a zone of inhibition does not appear, or is very small, then the bacterium is resistant to the antibiotic.
To make this aesthetic interpretation of the disc diffusion assay, I chose to use two naturally pigmented bacteria, Serratia marcescens (Sm), red, and Chromobacterium violaceum (Cv), purple and to test their sensitivity to three antibiotics, namely Cloxacillin (C), Kanamycin (K) and Trimethoprim (T). Moreover, instead of using the conventional circular discs for the assay and chose to use antibiotic impregnated papers that had been cut into letters.
In the first run of this process, the sensitivity of Sm and Cv to C and were investigated. After 12 hours incubation it became apparent that CV was resistant C, as it grew right up to the antibiotic impregnated paper, and in fact, even grew though it. On the other hand Sm was sensitive to C, as evidenced by the large and clear zone of inhibition (see below).
After 16 hours of incubation, mutant colonies of Sm that were resistant to C had begun to emerge. A direct visualisation of evolution and the emergence of antimicrobial resistance (see below).
After 24 hours incubation the C resistant mutants of Sm are clearly visible but neither bacterial strain, Sm or Cv, displays any resistance to K (see below).
In the second run of this process, the same bacteria were used, but the antibiotic Trimethoprim (T) replaced C and K. In the image below is the first run of this process (top) and run with T (bottom) but before incubation.
After 12 hours of incubation both Sm and Cv exhibit sensitivity to T, as evidenced by large and clear zones of inhibition(see below).
In a very unexpected development, after 24 hours of incubation Sm, but not Cv, had become resistant to T. However, Sm in doing so, had lost its ability to produce its red pigment and thus become white in the process (see below).